Tube: means capillary, tube, pipette tip, with any geometry of hole such as round, square, cylindrical, triangular, elliptical, parabolic or any shape which can be in the form of a tube. Internal diameter of the tube is from 0.001-25 mm. The length of the tube can be from 0.1-10000 mm.
Narrow Spiral Opening: means that the tube or capillary is twisted along the length of the tube in such a way that one end is fixed and other end (rotating end) is twisted with force at any given angle to form a spiral shape. The center of the spiral can be at any place between the two ends of the tube. During the twisting process, the length of the tube can be changed or unchanged depending on the application.
Spiral shape is created by twisting the tube at any angle from 1 degree to 360 degree or multiple turns of an angle between 1-360 degree.
Chromatographic material means, the regular, irregular, spherical, broken particles of silica, metal, polymers, metal oxides, non-metal oxides. These particles can be porous or nonporous. The pore size can be from 20-40000 nm. These particles can be modified chemically, physically, mechanically or by affinity media. The size of the particles can be from 0.001-1 mm.
Although a spectrum of analytical methods for small sample separation and purification have been developed, a number of problems, such as the slow speed of the separation process and the loss of sample volumes, limit the quality of currently available methods.
In this invention, we describe the use of a tube such as pipette tip, which has a spiral narrow opening at the lower end of the tip, designed for sample filtration or separation. The spiral narrow opening is of such dimensions that fluids and smaller particles can pass through the lower end while larger particles are retained. The application of such a device is useful for performing sample separation and filtration with chromatography and other separation media because it eliminates the need to place a filter or secondary solid matrices into the tip to retain said chromatography and other separation media within the tip during sample preparation. Depending on the specifications of the chromatography material, selected molecules from the sample can be separated or purified by binding to, or by being entrapped into the said material. The bound molecules can later be eluted from the said material by the use of different solvents.
Filters and secondary solid matrices are traditionally used in separation columns such as pipette tips to retain the solid matrix or column material in the container, while letting fluids and desired sample components flow through. Currently available sample preparation pipette tips rely on filters and solid matrices to hold chromatography and separation media in the tip while permitting selective components of the sample to pass through the lower end of the tip. The filters used in such devices may be made of silica, polypropylene, nylon, polytetrafluoroethylene or any other inert materials that do not react with the elution solution. The pore size of the filter material should be smaller than the particle size of the solid matrix or chromatography material placed above it so that the solid matrix or chromatography material is retained in the tip.
In pipette tips or other separation columns using filters, sample separation can be performed by centrifugation, gravitation, and vacuum suction, pressure application or by syringe- or pipette-based sample delivery through the tip or column. Such devices are used for the separation and purification of small sample volumes of bio-molecules such as proteins, peptides and DNA. The solid matrix or column material can consist of any material such as gel-filtration, affinity, ion-exchange, reverse-phase, and silica or modified-silica materials.
When sample volumes in the micro liter range are separated using currently available pipette tips and other separation/chromatography columns, one of the commonly associated problems is sample loss during the separation process due to retention of the sample on the filter or within the secondary solid matrices. Since the concentration of bio-molecules in micro volume samples is very small, the retention of molecules in the filter or secondary solid matrix can result in significant loss of the total sample volume. Also, since the volume of the filter or secondary solid matrix is often as large as the volume of the micro volume sample itself, the separation or chromatography process is adversely affected due to the large volume of filter material through which the sample must pass during the separation process. The filter material may also absorb proteins or bio-molecules from the sample, resulting in lower than desirable sample recovery.
Also, the filter material and solid matrix or column material may behave differently in different elution media, subsequently interfering with both the quality of the separation process and the volume of the sample retained. Furthermore, the presence of the filter material also slows down the sample separation process because the sample is first passed through the column material and then the filter prior to the completion of sample separation. Additionally, the structure and design of many micro-pipette tips, designed for micro-liter sample volumes, is not well suited for the addition of filters or secondary solid matrices into the tip.
The invention described and claimed herein comprises a tube or pipette tip designed to perform filtration or sample separation on the basis of a narrow spiral opening in the tip for sample filtration or separation. The narrow spiral opening is of such dimensions that fluids and smaller particles can pass through while larger particles are retained in the tip. During the process of sample separation, performed by centrifugation, gravitation, vacuum suction, pressure application or by syringe-based sample delivery through the container described herein, the filter solid matrix or column material will be retained in the tip while the sample will pass through.
The features of the present invention eliminate the need for filters or secondary solid matrices at the bottom of the tip to support the chromatography material during sample separation. Thus, the present invention, as described herein, will result in higher sample recovery due to the lack of sample retention on the filter or secondary solid matrix. The present invention will also eliminate separation problems associated with filter-materials and secondary solid matrices interacting differently with alternate elution media, thus improving the quality of the separation. The present invention also leads to more rapid sample separation since the sample does not flow through a filter or secondary solid matrices as part of the separation process.
The various features of novelty, which characterize the invention, are pointed out with particularity in the claims annexed to and forming a part of this disclosure. For a better understanding of the invention, its advantages and objects, reference is made to the accompanying drawings and descriptive matter in which a preferred embodiment of the invention is illustrated.